Major depressive disorder (MDD) is a recurring psychiatric disorder that affects up to 17% of the American population and can be severely debilitating by affecting sleep, work, social relationships, and appetite (Kessler et al., 2005). Although medications to relieve depressive symptoms exist, there is a time lag of weeks to months, and some patients receive no benefit from these drugs and are considered treatment-resistant. Both clinical and pre-clinical studies have revealed that prolonged negative stress can lead to MDD due to atrophy of neurons and related behavioral alterations; however, the molecular mechanisms underlying neuronal atrophy in response to chronic stress remain elusive, and an understanding of these mechanisms is critical to the development of new potential molecular targets for the pharmacological treatment of MDD. Thus, the present proposal is aimed at elucidating the molecular mechanisms by which stress causes atrophy of neurons in the PFC and subsequent depressive-like behaviors. Recent studies from our laboratory have pointed to a potential molecular mechanism for these effects. We have found that chronic unpredictable stress (CUS), an animal model for depression, decreases mTOR, a kinase that has been implicated in protein synthesis dependent synaptic plasticity. Consistent with this, a recent postmortem study reported that mTOR signaling is decreased in PFC of depressed subjects (Jernigan et al., 2011). Based in part on these findings, we hypothesize that stress can regulate changes at the synapse through the mTOR signaling pathway via RTP801, a stress- and glucocorticoid-induced protein and negative regulator of mTOR. In support of this hypothesis, we have shown that CUS increases RTP801 in the PFC, in parallel with a decrease in mTOR signaling and synaptic protein synthesis. Moreover, we have found in preliminary studies that mutant mice with a deletion of RTP801 are resistant to CUS. Accordingly, the experiments outlined in the present proposal seek to thoroughly test this hypothesis using a combination of genetic, behavioral, pharmacological, molecular, and biochemical techniques. In Specific Aim I, we will first characterize the role of RTP801 in mediating the behavioral, mTOR signaling, and synaptic changes following CUS. In Specific Aim II, we will directly test the hypothesis that the PFC is responsible for the behavioral changes induced by stress and regulated by RTP801, by local manipulation of RTP801 using viral vectors. This proposal fills a particularly critical area of research because of the numerous clinical implications that stem from understanding the molecular mechanisms of stress and the potential for identifying novel molecular targets for treating MDD.